DNA damage threatens the longevity and functionality of cells, and is common to the pathogenesis of many diseases. Better tools are needed for clinical identification and research into mechanisms underlying DNA oxidation, deamination, and hyper/hypomethylation, and the role these lesions play in the genesis and etiology of Alzheimer's disease. All existing and proposed techniques of DNA sequencing are insensitive to a broad spectrum of disease-relevant types of nucleobase damage Oxidative stress is one of the most earliest and prominent features of Alzheimer disease. Prior work of our collaborator and consultant, Dr. George Perry at University of Texas San Antonio and Dr. Xiongwei Zhu at Case Western Reserve University, demonstrated increased oxidative damage to nucleic acid in vulnerable neurons in the hippocampal tissues from Alzheimer patients. Due to the limits of technology available, it has not been possible to conclusively demonstrate oxidation in DNA is an upstream cause of AD and neuronal damage. It is also critical that vulnerable and influential genes, promoter, and sequences be identified to guide clinical prophylaxis and treatment. RMD Inc. is working to develop a label-free single-molecule-sensitive optical sequencing technology, capable of locating modified bases within single or double stranded DNA molecules in vitro. This technology will be capable of identifying and localizing genomic damage, while taking advantage of long read lengths and direct sequencing of unamplified DNA. In particular, this technology will use Raman spectroscopy with AFM (Tip-Enhanced Raman Spectroscopy) to directly read immobilized DNA for oxidized or otherwise non-canonical bases (i.e. methylated de-aminated, alkylated, etc.). The development of the new technology as proposed is a much needed one to further oxidative stress study in AD research. Our Phase I exploratory research will prove the principle of damage detection, and determine the initial resolution of the sequencing technique. Optics will be optimized, and the equipment will be evaluated for the capability to achieve single-base resolution in Phase II. In Phase II, RMD will achieve single base resolution, and apply the technology to the analysis of AD-derived DNA. The lab of Dr. Xiongwei Zhu will isolate pyramidal neurons in the CA1 area of hippocampus from well characterized AD patients and age-matched control patients by laser captured microdissection and extract DNA from these neurons and provide the DNA to RMD Inc. for further characterization of DNA oxidation. Experiments will be planned and directed by both Dr. George Perry and scientists at RMD.